Although great progress has been made in uncovering the genetic basis of many human cancers, it has not been possible so far to identify the molecular pathogenesis of hepatocellular carcinomas (HCC), a tumor of major public health importance worldwide. We have analyzed changes in specific genes in hepatocarcinogenesis, created models for the studies of cell proliferation, apoptosis and tumor development in genetically modified mice and demonstrated the key role of oxidative injury in hepatocyte apoptosis induced by diverse agents. We now propose to focus our studies on 2 general processes, which are nearly universal in liver carcinogenesis, namely, persistent cycles of hepatocyte apoptosis/proliferation and multiple chromosomal defects. We propose that cycles of hepatocyte apoptosis/regeneration result in constitutive hepatocyte replication and dysregulation of cell cycle checkpoints associated with defects in DNA double-strand breaks (DSB) repair mechanisms. Because of its major role in DSB repair and telomere capping, we will focus primarily, but not exclusively, on components of the non-homologous end-joining (NHEJ) DSB repair pathway. Using specially engineered mouse models and cell lines established in this laboratory, we have designed experiments to determine: a) whether repeated apoptosis/proliferation in mouse liver results in constitutive hepatocyte replication and chromosomal aberrations; b) if the high chromosomal instability in mouse liver carcinogenesis is associated with defects in DSB repair pathways; c) if hepatocarcinogenesis is enhanced in knockout mice deficient in components of DSB repair pathways exposed to the liver carcinogen diethylnitrosamine (DEN) or crossed with Transforming Growth Factor alpha (TGFalpha) transgenic mice, and d) whether growth abnormalities and chromosomal aberrations can be induced in normal and repair-deficient cultured hepatocytes transduced with TGFalpha and c-myc genes. We expect that these studies will contribute to the understanding of the molecular pathogenesis of HCC development in mice and humans.